Method of treating or ameliorating the symptoms of common cold or allergic rhinitis

ABSTRACT

This invention provides methods for the treatment or amelioration of the symptoms of the common cold or allergic rhinitis which comprises administering to a mammal in need thereof a 5-HT 2  antagonist.

This application claims priority under 35 USC 119 (e) over U.S.Application Ser. No. 60/014,038, filed Mar. 15, 1996.

FIELD OF THE INVENTION

The present invention is directed to the use of 5-HT2 antagonists fortreating or ameliorating the symptoms of the common cold or allergicrhinitis.

BACKGROUND OF THE INVENTION

Since the discovery of serotonin (5-hydroxytryptamine, 5-HT) over fourdecades ago, the cumulative results of many diverse studies haveindicated that serotonin plays a significant role in the functioning ofthe mammalian body, both in the central nervous-system and in peripheralsystems as well. Morphological studies of the central nervous systemhave shown that serotonergic neurons, which originate in the brain stem,form a very diffuse system that projects to most areas of the brain andspinal cord. R. A. O'Brien, Serotonin in Mental Abnormalities, 1:41(1978); H. W. M. Steinbusch, HANDBOOK OF CHEMICAL NEUROANATOMY, Volume3, Part II, 68 (1984); N. E. Anden, et al., Acta PhysiologicaScandinavia, 67:313 (1966). These studies have been complemented bybiochemical evidence that indicates large concentrations of 5-HT existin the brain and spinal cord. H. W. M. Steinbusch, supra.

With such a diffuse system, it is not surprising that 5-HT has beenimplicated as being involved in the expression of a number of behaviors,physiological responses, and diseases which originate in the centralnervous system. These include such diverse areas as sleeping, eating,perceiving pain, controlling body temperature, controlling bloodpressure, depression, schizophrenia, and other bodily states. R. W.Fuller, BIOLOGY OF SEROTONERGIC TRANSMISSION, 221 (1982); D. J. Boullin,SEROTONIN IN MENTAL ABNORMALITIES 1:316 (1978); J. Barchas, et al.,Serotonin and Behavior, (1973).

Serotonin plays an important role in peripheral systems as well. Forexample, approximately 90% of the body's serotonin is synthesized in thegastrointestinal system, and serotonin has been found to mediate avariety of contractile, secretory, and electrophysiologic effects inthis system. Serotonin may be taken up by the platelets and, uponplatelet aggregation, be released such that the cardiovascular systemprovides another example of a peripheral network that is very sensitiveto serotonin. Given the broad distribution of serotonin within the body,it is understandable that tremendous interest in drugs that affectserotonergic systems exists. In particular, receptor-specific agonistsand antagonists are of interest for the treatment of a wide range ofdisorders, including anxiety, depression, hypertension, migraine,compulsive disorders, schizophrenia, autism, neurodegenerativedisorders, such as Alzheimer's disease, Parkinsonism, and Huntington'schorea, and cancer chemotherapy-induced vomiting. M. D. Gershon, et al.,THE PERIPHERAL ACTIONS OF 5-HYDROXYTRYPTAMINE, 246 (1989); P. R. Saxena,et al., Journal of Cardiovascular Pharmacology, 15: Supplement 7 (1990).

Serotonin produces its effects on cellular physiology by binding tospecialized receptors on the cell surface. Multiple types of receptorsexist for many neurotransmitters and hormones, including serotonin. Theexistence of multiple, structurally distinct serotonin receptors hasprovided the possibility that subtype-selective pharmacologic agents canbe produced. The development of such compounds could result in new andincreasingly selective therapeutic agents with fewer side effects, sinceactivation of individual receptor subtypes may function to affectspecific actions of the different parts of the central and/or peripheralserotonergic systems.

An example of such specificity can be demonstrated by using the vascularsystem as an example. In certain blood vessels, stimulation of certain5-HT receptors on the endothelial cells produces vasodilation whilestimulation of certain 5-HT receptors on the smooth muscle cellsproduces vasoconstriction.

Currently, the major classes of serotonin receptors (5-HT₁, 5-HT₂,5-HT₃, 5-HT₄, 5-HT₅, 5-HT₆, and 5-HT₇) contain some fourteen to eighteenseparate receptors that have been formally classified based on theirpharmacological or structural differences. For an excellent review ofthe pharmacological effects and clinical implications of the various5-HT receptor types, see Glennon, et al., Neuroscience and BehavioralReviews, 14:35 (1990).! discoveries.

One class of serotonin receptors is the 5-HT₂. Of this class, severalsubtypes are known to exist. These subtypes include 5-HT_(2A), 5-HT_(2B)and 5-HT_(2C). The subtype 5-HT_(2A) is located in the vascular smoothmuscle, platelets, lung, CNS and gastrointestinal tract. This receptoris thought to be associated with vasoconstriction, platelet aggregation,and bronchoconstriction. The 5-HT_(2B) receptor is localized in the ratlung, stomach fundus, uterus, bladder, and colon. Interesting areas of5-HT_(2B) receptor localization in the human include, but are notlimited to, the brain and blood vessels. Subtype 5-HT_(2C) is located inthe CNS with a high density in the choroid plexus.

Pollen has long been recognized as a cause of allergic rhinitis commonlycalled "hay fever". Pollen contains proteases which induce the releaseof mediators from mast cells, thereby stimulating IgE biosynthesis. Thedegranulation of mast cells by IgE results in the release of histaminewhich leads to an inflammatory response which causes congestion,itching, and swelling of sinuses. Degranulation of mast cells can alsobe caused by inflammatory neuropeptides, like substance P, released fromsensory nerves. The histamine released by mast cells followingdegranulation can activate sensory nerves increasing the release ofinflammatory neuropeptides and sending signals to the brain causingsneezing and the spread of the allergic reaction to other, nearby areas,for example, causing watery eyes. In this way, sensory nervesparticipate in the inflammation process. Many eosinophils are present inallergic patients with nasal mucus and neutrophils are present inpatients with infected mucus.

Antihistamines are drugs commonly utilized which, when taken orally,frequently have a sedative effect. Alternatively, nasal sprayscontaining cromolyn sodium have been effective as cromolyn acts byblocking the reaction of the allergen with tissue mast cells. Cromolynis not entirely effective, however, as it apparently does not bind tosome of the mediators of inflammation or the activators of IgEbiosynthesis that stimulate the degranulation of mast cells and theproduction of histamine from the mast cells.

Inflammation is a non-specific response of tissues to diverse stimuli orinsults and results in the release of materials at the site ofinflammation that induce pain. It is now recognized that mast cells,neutrophils, T-cells and sensory nerves are implicated in thepathophysiology of inflammatory skin conditions as well as in otherphysiological disorders. Mast cells provide the greatest source ofhistamines in acute inflammation, as well as chymases, afterdegranulation by IgE.

The "common cold" is a time honored phrase used by both physicians andlay persons alike for the identification of acute minor respiratoryillness. Since the identification of rhinovirus in 1956, a considerablebody of knowledge has been acquired on the etiology and epidemiology ofcommon colds. It is known that the common cold is not a single entity,but rather is a group of diseases caused by members of several familiesof viruses, including parainfluenza viruses, rhinoviruses, respiratorysyncytial viruses, enteroviruses, and coronaviruses. Much work has beenperformed in characterizing viruses which cause the common cold. Inaddition, the molecular biology of rhinoviruses, the most importantcommon cold viruses, is understood in great detail. In contrast,progress on the treatment of common colds has been slow despite theseadvances. While there are now large numbers of compounds that have beenfound to exhibit antiviral activity against cold viruses in cellculture, antiviral compounds have had limited effectiveness in patients.

Because of the widespread dissatisfaction with the currently marketedtreatments for the common cold and allergic rhinitis within the affectedpopulation, there exists a need for a more efficacious and safetreatment. The present invention provides such a treatment.

SUMMARY OF THE INVENTION

This invention provides a method for the treatment or amelioration ofthe symptoms of the common cold or allergic rhinitis in a mammal whichcomprises administering to a mammal in need thereof an effective amountof a compound having activity as a 5-HT₂ antagonist.

DEFINITIONS

The terms and abbreviations used in the instant preparations andexamples have their normal meanings unless otherwise designated. Forexample "°C." refers to degrees Celsius; "N" refers to normal ornormality; "mmol" refers to millimole or millimoles; "g" refers to gramor grams; "ml" means milliliter or milliliters; "L" means liter orliters; "M" refers to molar or molarity; "MS" refers to massspectrometry; "IR" refers to infrared spectroscopy; and "NMR" refers tonuclear magnetic resonance spectroscopy.

The term "allergic rhinitis" as employed herein is understood to includerhinitis medicamentosa, rhinitis sicca, and atrophic rhinitis.

DETAILED DESCRIPTION AND PREFERRED EMBODIMENTS

The method of the present invention employs 5-HT₂ receptors. There arethree members of the 5-hydroxytryptamine 2 (5-HT₂) family of 5-HTreceptors, 5-HT_(2A), 5-HT_(2B) and 5-HT_(2C) receptors. These receptorsare G-protein linked receptors that are positively coupled tophosphoinositide metabolism, at least in the cloned versions of thesereceptors. These receptors share sequence homology and have the samepattern of introns and exons. Similarities in the specificity of thereceptors for the ligands further indicates the commonality of receptorsin this family. While the method of the present invention can employ anyof the 5-HT₂ receptor subtypes, a more preferred receptor subtype is5-HT_(2B).

The present invention provides a method for the treatment oramelioration of the symptoms of the common cold or allergic rhinitiswhich comprises administering to a mammal in need thereof an effectiveamount of a 5-HT₂ receptor antagonist. A more preferred embodiment ofthe present invention provides a method for the treatment oramelioration of the symptoms of the common cold or allergic rhinitiswhich comprises administering to a mammal in need thereof an effectiveamount of a 5-HT_(2B) receptor antagonist.

In recent publications many different 5-HT₂ receptor antagonists whichcan be utilized in the present method have been described.

For instance, U.S. Pat. No. 5,428,036, incorporated herein by reference,describes a group of 5-HT₂ antagonists of Formula II: ##STR1## wherein Xis selected from CO, CS or CH₂, and if X is CO or CS, R is selected fromthe group consisting of:

i) hydrogen, C₁ -C₂₄ alkyl, C₂ -C₂₄ alkenyl, C₃ -C₈ cycloalkyl, C₃ -C₈cycloalkenyl or C₄ -C₃₂ cycloalk(en)ylalk(en)yl, optionally substitutedwith one or two hydroxy groups, or phenyl optionally substituted withone or more substituents selected from the group consisting of halogen,trifluoromethyl, C₁ -C₄ alkyl, C₁ -C₄ alkoxy, C₁ -C₄ alkylthio, acyloxy,or cyano; or

ii) YR¹, wherein Y is O or S and R¹ is selected from the substituentsdefined for R under i) above; and

iii) NR² R³, wherein R² and R³ independently are selected from thesubstituents defined for R under i) above or R² and R³ are combined toform a four to eight member heterocyclic ring containing from one tothree nitrogen atoms and from zero to three oxygen or sulfur atoms; orif X is CH2, R is selected from the groups consisting of:

iv) a group YR¹ as defined in ii);

v) a group NR² R³ as defined in iii); or

vi) a group OC(O)R⁴, wherein R⁴ is as defined for R¹ ; andpharmaceutically acceptable salts thereof.

Another group of 5-HT₂ antagonists include the compounds described inU.S. Pat. No. 5,229,382, incorporated herein by reference, which are ofthe general Formula III: ##STR2##

Still another group of 5-HT₂ antagonists are those in U.S. Pat. No.5,457,115, incorporated herein by reference, which describes antagonistsof the Formula IV: ##STR3## where Ar is one of a phenyl group, a phenylgroup substituted with at least one substituent selected from halogen,lower alkyl, lower alkoxy, hydroxy, trifluoromethyl, and cyano, and ahetero aromatic group selected from 2-thienyl, 3-thienyl, 2-furanyl,3-furanyl, 2-oxazolyl, 2-imidazolyl, 2-pyridyl, 3-pyridyl, and4-pyridyl; each dotted line is an optional double bond; X and X¹ areindependently selected from the group consisting of hydrogen, halogen,lower alkyl, lower alkoxy, hydroxy, lower alkylthio, loweralkylsulfonyl, lower alkylamino, lower dialkylamino, cyano,trifluoromethyl, and trifluoromethylthio; or X and X¹ are taken togetherto form a 5 to 7 membered carbocyclic ring; R¹ is selected from thegroup consisting of hydrogen, lower alkyl and alkyl substituted with oneor two hydroxy groups; with the proviso that when X is hydrogen orfluoro then R¹ cannot be hydrogen; R is a substituent having theformula: ##STR4## wherein n is an integer from 2-6 inclusive; W isoxygen or sulfur; V¹ is selected from OR⁴, SR⁵, CHR⁶ R⁷, and NR⁸ R⁹ ;

wherein R³ to R⁹ are independently selected from the group consisting ofhydrogen, lower alkyl, lower alkenyl, cycloalkyl, lower alkylsubstituted with one or two hydroxyl groups; and lower alkenylsubstituted with one or two hydroxyl groups; and pharmaceuticallyacceptable acid addition salts or prodrugs thereof.

An even further group of 5-HT₂ antagonists which can be utilized in thepresent method include those in U.S. Pat. No. 5,480,885, incorporatedherein by reference, which describes antagonists of the Formula V:##STR5## wherein R₁, R₂ and R₃ independently represent a hydrogen atomor a straight-chain or branched-chain C₁ -C₆ alkyl group,

X represents a hydrogen or a halogen atom;

Z represents a carbonyl or methylene group and C₉ --C₁₀ represents asingle or a double bond, racemates and acid addition salts thereof.

The above groups of compounds are only illustrative of the 5-HT₂receptor antagonists which are currently under development. This listingof groups of compounds is not meant to be comprehensive, the methods ofthe present invention may employ any 5-HT₂ receptor antagonist and isnot limited to any particular class of compound.

A more preferred class of antagonists are the 5-HT₂ receptor antagonistsof Formula VI: ##STR6## wherein R¹ is hydrogen or C₁ -C₃ alkyl;

R₃ is hydrogen or C₁ -C₃ alkyl;

R⁶ is selected from the group consisting of hydrogen, C₁ -C₆ alkyl, C₂-C₆ alkenyl, halo, halo(C₁ -C₆)alkyl, halo(C₂ -C₆)alkenyl, COR⁵, C₁ -C₁₀alkanoyl, CO₂ R^(5'), (C₁ -C₆ alkyl)_(m) amino, NO₂, --SR⁵, and OR⁵ ;

R⁷ and R⁸ are independently selected from the group consisting ofhydrogen, C₁ -C₆ alkyl, C₂ -C₆ alkenyl, NO₂, halo, halo(C₁ -C₆)alkyl,halo(C₂ -C₆)alkenyl, COR⁵, C₁ -C₁₀ alkanoyl, C₇ -C₁₆ arylalkyl, CO₂R^(5'), (C₁ -C₆ alkyl)_(m) amino, --SR⁵, and OR⁵ ;

n is 1, 2, or 3;

n' is 1, 2, or 3;

m is 1 or 2;

R⁵ is independently hydrogen or C₁ -C₄ alkyl;

R^(5') is C₁ -C₄ alkyl;

-- is optionally a bond;

a pharmaceutically acceptable salt or solvate thereof.

Examples of compounds of Formula VI include but are not limited to:spiro-9,92-(3,4-dichloro)-1,2,3,4-tetrahydronaphthyl!-5-methoxy-1,2,3,9-tetrahydro-8H-pyridoindole, spiro-9,92-(3,4-dimethoxy)-1,2,3,4-tetrahydronaphthyl!-5-methyl-1,2,3,9-tetrahydro-8H-pyridoindole, spiro-9,92-(3,4-diethoxy)-1,2,3,4-tetrahydronaphthyl!-5-methyl-1,2,3,9-tetrahydro-8H-pyridoindole, spiro-9,92-(3,5-dichloro)-1,2,3,4-tetrahydronaphthyl!-5-dimethylamino-1,2,3,9-tetrahydro-8H-pyridoindole, spiro-9,92-(3-fluoro,4-chloro)-1,2,3,4-tetrahydronaphthyl!-5-ethyl-1,2,3,9-tetrahydro-8H-pyridoindole, spiro-9,9 2-(3,4-dimethoxy)-1,2,3,4-indole, spiro-9,92-(3,4-dimethoxy)-1,2,3,4-tetrahydronaphthyl!-5-bromo-1,2,3,9-tetrahydro-8H-pyridoindole, spiro-9,92-(3,4-dimethoxy)-1,2,3,4-tetrahydronaphthyl!-5-chloro-1,2,3,9-tetrahydro-8H-pyridoindole.

The synthesis of these compounds is described in co-pending UnitedStates Provisional Patent Application Ser. No. 06/014,119, AttorneyDocket No. P-10656, filed Mar. 25, 1996, incorporated herein byreference. The syntheses of typical compounds from this class, includingsix specific examples, are detailed infra.

The compounds of Formula VI can be prepared using chemical processesthat are understood in the art. The examples are illustrative only, andare not intended to limit the scope of the invention.

Indole Starting Materials

The indole starting materials (1a, 1b, and 1c) infra. were purchased(1a), prepared according to Bartoli's procedure (1b) G. Bartoli, et al.,Tetrahedron Lett., 1989, 30, 2129! or (1c) synthesized from2-Iodo-4,6-dimethylaniline (5'"). The process is illustrated by thefollowing Scheme: ##STR7##

The 2-Iodo-4,6-dimethylaniline (5"') synthesis can be completed asfollows: To a suspension of 5"' (24 mmol.), CuI (0.05 equiv.) and(PPh₃)₂ PdCl₂ (0.05 equiv.) in 30 ml of dry triethylamine under Ar.atmosphere was added trimethylsilylacetylene (1.1 equiv.) and theresulting mixture was stirred for 3 hours. Then, the solvent waseliminated under vacuum and the residue purified by flash chromatographyusing hexane/ethyl acetate (3:1) as eluent to yield 6" in quantitativeyield. A slurry of 6"' (23 mmol.) and CuI (2 equiv.) in 50 ml of drydimethyl formamide was heated for 2.5 h. under Ar. atmosphere at 100° C.After cooling down to room temperature the reaction mixture was filteredoff and the solid washed twice with ether (20 ml.). The organic phasewas washed with water (3×50 ml.), dried over Na₂ SO₄ and the solventevaporated to dryness. The crude product was purified by flashchromatography using hexane/ethyl acetate (3:1) as eluent to afford 1c(1.5 g., 45%).

EXAMPLE 1 ##STR8##

A suspension of the corresponding tryptamine hydrochloride (3a) (1 gram)and the corresponding dimethoxytetralone (3b) (1 gram) in ethanol (10ml.) was refluxed during 128 h. After this time the reaction mixture wasallowed to reach room temperature and filtered off. The crude solid waswashed and dried. Melting point 261° C.

    ______________________________________                 Theory                       Found    ______________________________________    C              69.25   69.34    H              6.82    6.97    N              7.02    6.98    ______________________________________

EXAMPLE 2 ##STR9##

A suspension of the corresponding tryptamine hydrochloride (2a) (575 mg)and the corresponding ketone (2b) (464 mg) in ethanol (10 ml.) wasrefluxed during 128 h. After this time the reaction mixture was allowedto reach room temperature and filtered off. The crude solid was washedand dried.

Yield: 525 mg

    ______________________________________                 Theory                       Found    ______________________________________    C              74.43   74.36    H              6.84    6.84    N              8.27    8.25    ______________________________________

MS: 301

EXAMPLE 3 ##STR10##

A suspension of the corresponding tryptamine hydrochloride (2a) (500 mg)and the corresponding ketone (2b) (396 mg) in ethanol (10 ml.) wasrefluxed during 72 h. After this time the reaction mixture was cooled toabout 0° C. and filtered off. The crude solid was washed and dried.

Yield: 262 mg

MS: 274

EXAMPLE 4 ##STR11##

A suspension of the corresponding tryptamine hydrochloride (4a) (500 mg)and the corresponding ketone (4b) (396 mg) in ethanol (10 ml.) wasrefluxed during 72 h. After this time the reaction mixture was cooled toabout 0° C. for about 24 hours and filtered off. The crude solid waswashed and dried.

Submitted for mass spectral analysis and found mi of 274.

EXAMPLE 5 ##STR12##

A suspension of the corresponding tryptamine hydrochloride (5a) (500 mg)and the corresponding ketone (5b) (397 μL) in ethanol (10 ml.) wasrefluxed during 72 h. After this time the reaction mixture was cooled toabout 0° C. for 14 hours and filtered off. The crude solid was washedand dried.

Yield: 630 mg

    ______________________________________                 Theory                       Found    ______________________________________    C              73.95   73.32    H              6.52    6.73    N              8.62    8.59    ______________________________________

MS: 288

EXAMPLE 6 ##STR13##

A suspension of the corresponding tryptamine hydrochloride (4a) (1 g)and the corresponding ketone (4b) (800 mg) in ethanol (10 ml.) wasrefluxed during 72 h. After this time the reaction mixture was cooled toabout 0° C. for about 24 hours and filtered off. The crude solid waswashed and dried.

Yield: 550 mg

    ______________________________________                 Theory                       Found    ______________________________________    C              70.67   70.88    H              7.06    7.16    N              7.85    7.88    ______________________________________

An additional preferred class of 5-HT₂ receptor antagonists are thosecompounds described in WO 95/24200, incorporated herein by reference,which are of Formula I: ##STR14## wherein Q is hydrogen or (CHR₂)R₄ ;

R₁ is hydrogen or C₁ -C₃ alkyl;

R₂ is hydrogen or C₁ -C₃ alkyl;

R₃ is hydrogen or C₁ -C₃ alkyl;

R₄ is C₅ -C₈ cycloalkyl, substituted C₅ -C₈ cycloalkyl, C₅ -C₈cycloalkenyl, substituted C₅ -C₈ cycloalkenyl, bicyclic or substitutedbicyclic;

A is selected from the group consisting of ##STR15## wherein, R₆ and R₇are, independently, hydrogen, C₁ -C₆ alkyl, C₂ -C₆ alkenyl, halo,halo(C₁ -C₆) alkyl, halo(C₂ -C₆)alkenyl, COR₅, C₁ -C₁₀ alkanoyl, CO₂R_(5'), (C₁ -C₆ alkyl)_(m) amino, NO₂, --SR₅, or OR₅ ;

m is 1 or 2;

R₅ is independently hydrogen or C₁ -C₄ alkyl;

R₅, is C₁ -C₄ alkyl;

R₈ is independently selected from the group consisting of an R₆ group,substituted C₃ -C₈ cycloalkyl, C₃ -C₈ cycloalkyl, C₃ -C₈ cycloalkyl-(C₁-C₃)alkyl, C₅ -C₈ cycloalkenyl, substituted C₅ -C₈ cycloakenyl, C₅ -C₈cycloalkenyl-(C₁ -C₃)alkyl, C₇ -C₁₆ arylalkyl; or

R₆ and R₇ together with the carbon atoms of group A form a 5- to8-member carbon ring; or a pharmaceutically acceptable salt or solvatethereof.

Examples of compounds of Formula I include but are not limited to:8-methyl-1- 3,4-dimethoxyphenyl)methyl!1,2,3,4-tetrahydro-9H-pyrido3,4-b!indole, 8-bromo-1-3,4-dimethoxyphenyl)methyl!-1,2,3,4-tetrahydro-9H-pyrido 3,4-b!indolehydrochloride, 6,8-dibromo-1-3,4-dimethoxyphenyl)methyl!-1,2,3,4-tetrahydro-9H-pyrido 3,4-b!indole;6-methyl-8-bromo-1-3,4-dimethoxyphenyl)-methyl!-1,2,3,4-tetrahydro-9H-pyrido 3,4-b!indolehydrochloride; 8-methoxy-1-3,4-dimethoxyphenyl)methyl!-1,2,3,4-tetrahydro-9H-pyrido 3,4-b!indole;6,8-difluoro-1-(3,4-dimethoxyphenyl)-methyl!-1,2,3,4-tetrahydro-9H-pyrido 3,4-b!indole;7-methyl-8-bromo-1-3,4-dimethoxyphenyl)-methyl!-1,2,3,4-tetrahydro-9H-pyrido 3,4-b!indolehydrochloride; 6-(1,1-dimethylethyl)-1-(3,4-dimethoxyphenyl)methyl!1,2,3,4-tetrahydro-1-9H-pyrido- 3,4b!indolehydrochloride; 5-fluoro-6-methyl-1-(3,4-dimethoxyphenyl)-methyl!-1,2,3,4-tetrahydro-9H-pyrido 3,4-b!indole;7,8,9,10-tetrahydro-10- (3,4-dimethoxyphenyl) methyl!-11H-benzo g!pyrido3,4-b!indole; 6-cyclohexyl-1-(3,4-dimethoxyphenyl)-methyl!-1,2,3,4-tetrahydro-9H-pyrido 3,4-b!indolehydrochloride; 5,8-dimethyl-1,2,3,4-tetrahydro-1-(3,4-dimethoxyphenyl)methyl!-9H-pyrido3,4b!indole hydrochloride;6-(1-methylethyl)-1,2,3,4-tetrahydro-1-(3,4-dimethoxyphenyl)methyl!-9H-pyrido 3,4b!indole;6,8-dimethyl-1,2,3,4-tetrahydro-1-(3,4-dimethoxyphenyl)methyl!-9H-pyrido3,4b!indole hydrochloride;5,7-dimethyl-1,2,3,4-tetrahydro-1-(3,4-dimethoxyphenyl)methyl!-9H-pyrido3,4b!indole hydrochloride;6,7-dimethyl-1,2,3,4-tetrahydro-1-(3,4-dimethoxyphenyl)methyl!-9H-pyrido3,4b!indole;6-ethyl-1,2,3,4-tetrahydro-1- (3,4-dimethoxyphenyl)methyl!-9H-pyrido3,4b!indole; 6-bromo-1,2,3,4-tetrahydro-1-(3,4-dimethoxyphenyl)methyl!-9H-pyrido 3,4b!indole;7,8-dimethyl-1,2,3,4-tetrahydro-1-(3,4-dimethoxyphenyl)methyl!-9H-pyrido3,4b!indole hydrochloride;6-methyl-1,2,3,4-tetrahydro-1- (3,4-dimethoxyphenyl)methyl!-9H-pyrido3,4b!indole hydrochloride; 6-methyl-1-(3,4,5-trimethoxyphenyl)-methyl!-1,2,3,4-tetrahydro-9H-pyrido3,4-b!indolehydrochloride; 6-methyl-1-(2,3,4-trimethoxyphenyl)-methyl!-1,2,3,4-tetrahydro-9H-pyrido3,4-b!indolehydrochloride; 6-methyl-1-(2-methoxyphenyl)methyl!-1,2,3,4-tetrahydro-9H-pyrido 3,4-b!indolehydrochloride; 6-methyl-1-(2,4-dimethoxyphenyl)methyl!-1,2,3,4-tetrahydro-9H-pyrido 3,4-b!indolehydrochloride; 6-methyl-1-(2,5-dimethoxyphenyl)methyl!-1,2,3,4-tetrahydro-9H-pyrido 3,4-b!indolehydrochloride; 6-methyl-1-(2,4,5-trimethoxyphenyl)-methyl!-1,2,3,4-tetrahydro-9H-pyrido3,4-b!indolehydrochloride; 6-(1-methylethyl)-1-(2,3,4-trimethoxy-phenyl)methyl!-1,2,3,4-tetrahydro-9H-pyrido3,4-b!indole hydrochloride; 6-methyl-1-(3,4-dimethoxy-5-nitrophenyl)-methyl!-1,2,3,4-tetrahydro-9H-pyrido3,4-b!indole hydrochloride; 6-methyl-1-(3-iodo-4,5-dimethoxy-phenyl)-methyl!-1,2,3,4-tetrahydro-9H-pyrido3,4-b!indole; 6-methyl-1-(3,4-dimethoxy-5-amino-phenyl)-methyl!-1,2,3,4-tetrahydro-9H-pyrido3,4-b!indole dihydrochloride; 6-methyl-1-(3-methoxy-4-propoxyphenyl)-methyl!-1,2,3,4-tetrahydro-9H-pyrido3,4-b!indole; 6-methyl-1-(4-dimethylaminophenyl)methyl!-1,2,3,4-tetrahydro-9H-pyrido 3,4-b!indoledihydrochloride; 6-methyl-1-(4-dibutylaminophenyl)methyl!-1,2,3,4-tetrahydro-9H-pyrido 3,4-b!indoledihydrochloride; 6-methyl-1-(3-fluoro-4-methoxyphenyl)-methyl!-1,2,3,4-tetrahydro-9H-pyrido3,4-b!indole hydrochloride; 6-(1-methyl-1-(3,4-dimethylphenyl)methyl!-1,2,3,4-tetrahydro-9H-pyrido 3,4-b!indolehydrochloride; 6-methyl-1-(2-chloro-3,4-dimethoxyphenyl)-methyl!-1,2,3,4-tetrahydro-9H-pyrido3,4-b!indole hydrochloride; 6-methyl-1-(2-chloro-3-methoxy-4-hydroxyphenyl)methyl!-1,2,3,4-tetrahydro-9H-pyrido3,4-b!-indole hydrochloride; 5-fluoro-6-methyl-1-(2-chloro-3,4-dimethoxyphenyl)methyl!-1,2,3,4-tetrahydro-9H-pyrido3,4-b!indole hydrochloride;6-methyl-1-(cyclohexylmethyl)-1,2,3,4-tetrahydro-9H-pyrido 3,4-b!indolehydrochloride; 6-methyl-1-(2-bromo-3,4-dimethoxyphenyl)-methyl!-1,2,3,4-tetrahydro-9H-pyrido2,4-b!indole; and 6-iodo-1-(3,4-dimethoxyphenyl)-methyl!-1,2,3,4-tetrahydro-9H-pyrido 3,4-b!indolehydrochloride.

EXAMPLE 7 Preparation of 6-methyl-1-(2-chloro-3,4-dimethoxyphenyl)-methyl!-1,2,3,4-tetrahydro-9H-pyrido3,4-b!indole hydrochloride ##STR16##

A solution of 2-chloro-3,4-dimethoxybenzaldehyde (10.45 g),N-acetylglycine (11.9 g, 0.10 mol.) and sodium acetate (8.4 g, 0.1 mol)in acetic anhydride (100 mL) was heated to 100° C. for 2 hours. Thereaction mixture was cooled to ambient temperature poured onto ice (300mL) with stirring. The product was isolated by filtration, washed withwater (3×50 mL) and diethyl ether (3×50 mL) and dried under reducedpressure (5.26 g). ##STR17##

A suspension of azalactone prepared above (1.34 g, 4.76 mmol.) and5-methyltryptamine hydrochloride (1.0 g, 4.75 mmol.) in 1N HCl (30 mL)was heated to reflux for 24 hours under nitrogen atmosphere. Thereaction mixture was cooled to ambient temperature and the crude productisolated by filtration. The solid was triturated with ethanol and washedwith diethyl ether. The product was isolated by filtration (1.19 g).m/e=370, mp. 244° C. (dec.).

    ______________________________________    Analysis       Calculated                            Found    ______________________________________    C              61.92    61.67    H              5.94     5.94    N              6.88     6.94    ______________________________________

EXAMPLE 8 Preparation of 6-methyl-1-(2-bromo-3,4-dimethoxyphenyl)-methyl!-1,2,3,4-tetrahydro-9H-pyrido3,4-b!indole

Example 8 was prepared in the same manner as described in Example 7 withthe following exception: 2-bromo-3,4-dimethoxybenzaldehyde was used asstarting material instead of 2-chloro-3,4-dimethoxybenzaldehyde. Thefinal compound ##STR18## had a yield of 79.2%; M/I 416, 414; and mp272°-4° C.

    ______________________________________    Analysis       Calculated                            Found    ______________________________________    C              55.83    55.57    H              5.35     5.36    N              6.20     6.09    ______________________________________

EXAMPLE 9 Preparation of 6-iodo-1-(3,4-dimethoxyphenyl)-methyl!-1,2,3,4-tetrahydro-9H-pyrido 3,4-b!indolehydrochloride

Example 9 was prepared in the same manner as Example 7 with thefollowing exceptions: 3,4-dimethoxybenzaldehyde was used instead of2-chloro-3,4-dimethoxybenzaldehyde and 5-iodo-tryptamine instead of5-methyl-tryptamine as starting materials. Upon completion of thereaction, the mixture was neutralized with aqueous potassium carbonatesolution and extracted with chloroform. The combined chloroform phaseswere dried over anhydrous sodium carbonate and concentrated underreduced pressure. The product was purified by chromatography on silicagel, eluting with 2% methanol in chloroform. Fractions containingproduct were pooled and concentrated. The residue was dissolved indiethyl ether and was treated with gaseous HCl. The resulting HCl saltwas isolated by filtration and dried under reduced pressure. The finalcompound ##STR19## had a yield of 31.3%; M/I 448; and mp 270°-3° C.

    ______________________________________    Analysis        Calculated                             Found    ______________________________________    C               49.55    49.62    H               4.57     4.51    N               5.78     5.66    ______________________________________

The biological efficacy of a compound believed to be effective as a5-HT₂ receptor antagonist was confirmed by first employing an initialscreening assay which rapidly and accurately measures the binding of thetest compound to the 5-HT₂ receptor. Once the binding of the testcompound is established, the in vivo activity of the test compound onthe receptor is established. Assays useful for evaluating 5-HT₂antagonist are well known by those skilled in the art. Assays for eachof the 5-HT₂ receptors are included below.

5-HT_(2B) Receptor Binding Activity

The ability of a compound to bind to a 5-HT_(2B) receptor was measuredusing standard procedures such as that listed below.

Assay Procedure.

Certain compounds and intermediates of the present invention are usefulfor modulating 5-HT_(2B) receptors. The compounds which are most usefulfor binding a 5-HT_(2B) receptor can be identified using the followingprocedures. Further, a useful in vivo model for demonstrating 5-HT_(2B)activity is provided infra.

Radioligand Binding Studies for 5-HT_(2B) ; Membrane preparation fromtransformed cells

Suspension cells expressing the cloned rat 5-HT_(2B) receptor wereharvested by centrifugation at 2,200×g for 15 min at 4° C. J. D. Kursar,et al, Mol. Pharmacol., 42:549-557 (1992). Membranes for the bindingassays were prepared by vortexing the pellet in 50 mM Tris-HCl, pH 7.4(0.5×10⁹ cells/30 ml). The tissue suspension was then centrifuged at39,800×g for 10 min at 4° C. This procedure was repeated for a total ofthree washes, with a 10 minute incubation at 37° C. between the firstand second wash. The final pellet was homogenized in 67 mM Tris-HCl, pH7.4 (at 20-40 and 12.5 million cells/ml, original cell number, for cellsexpressing low and relatively high levels of the 5-HT_(2B) receptor,respectively) using a Tissumizer (Tekmar, Cincinnati, Ohio), setting 65for 15 seconds.

³ H!5-HT binding studies

Binding assays were automated using a Biomek 1000 (Beckman Instruments,Fullerton, Calif.) and were performed in triplicate in 0.8 ml totalvolume. Membrane suspension, 200 μl, (0.04-0.27 mg protein) and 200 μlof drug dilution in water were added to 400 μl of 67 mM Tris-HCl, pH7.4, containing ³ H!5-HT, pargyline, CaCl₂, and L-ascorbic acid. Finalconcentrations of pargyline, CaCl₂ and L-ascorbic acid were 10 μM, 3 mMand 0.1%, respectively. Tubes were incubated at 37° C. for 15 min or at0° C. for 2 hours (binding equilibria were verified for both of theseconditions), then rapidly filtered using a Brandel cell harvester (ModelMB-48R; Brandel, Gaithersburg, Md.) through Whatman GF/B filters whichhad been presoaked in 0.5% polyethylenimine and precooled with ice-cold50 mM Tris-HCl, pH 7.4. The filters were then washed rapidly four timeswith one ml ice-cold 50 mM Tris-HCl, pH 7.4. The amount of ³ H!5-HTtrapped on the filters was determined by liquid scintillationspectrometry (Ready Protein and Beckman LS 6000IC, Beckman Instruments,Fullerton, Calif.). For the saturation experiments, actual freeradioligand concentrations were determined by sampling the supernatantof parallel saturation experiments in which bound radioactivity had beenseparated by centrifugation. The concentration of ³ H!5-HT ranged from0.02 to 5 nM and 0.6 to 63 nM for saturation experiments incubated at 0°C. and 37° C., respectively. 5-HT, 10 μM, or 1-naphthylpiperazine(1-NP), 10 μM, defined nonspecific binding. For competition experiments,six to twelve concentrations of displacing drugs were used, spanning sixlog units, and the final concentration of ³ H!5-HT was 2 nM. Protein wasdetermined by the method of Bradford, using bovine serum albumin as thestandard. M. M. Bradford, Anal. Biochem., 72:248-254 (1976).

Statistical Analysis

The K_(d) and B_(max) values from the saturation assays were determinedfor best fit to a one-site or a two-site binding model using a partialF-test. A. De Lean, et al, Mol. Pharmacol., 21:5-16 (1981). Thefollowing equation was used for a one-site binding model, ##EQU1## whereBound=amount of ³ H!5-HT specifically bound, B_(max) =maximum number ofbinding sites, K_(d) =equilibrium dissociation constant and L!=freeconcentration of ³ H!5-HT, or a two-site binding model, ##EQU2## whereBound=amount of ³ H!5-HT specifically bound, B_(max) =maximum number ofhigh affinity binding sites, B_(max2) =maximum number of low affinitybinding sites, K_(d1) =equilibrium dissociation constant for the highaffinity site, K_(d2) =equilibrium dissociation constant for the lowaffinity site and L!=free concentration of ³ H!5-HT. The IC₅₀ valuesfrom the competition assays, the binding parameters for the IP₃ standardcurve and the EC₅₀ and E_(max) values from the IP₃ assays weredetermined by nonlinear regression analysis of four parameter logisticequations (Systat, Systat Inc, Evanston, Ill.). A. De Lean, et al, Mol.Pharmacol., 21:5-16 (1981). The IC₅₀ values were converted to K_(i)values using the Cheng-Prusoff equation. Y. Cheng, et al, Biochem.Pharmacol., 22:3099-3108 (1973).

Assay Methods 5-HT_(2B) in vitro

Male Wistar rats (150-375 g; Laboratory Supply, Indianapolis, Ind.) weresacrificed by cervical dislocation, and longitudinal section of thestomach fundus were prepared for in vitro examination. Four preparationswere obtained from one rat fundus. Ring preparations of the extractedjugular vein were prepared as described by Hooker; Blood Vessels, 14:1(1977) and M. L. Cohen, J. Pharmacol. Exp. Ther. 227:327 (1983). Tissueswere mounted in organ baths containing 10 mL of modified Krebs solutionof the following composition (millimolar concentrations): NaCl, 118.2,KCl, 4.6; CaCl₂.H₂ O, 1.6; KH₂ PO₄, 1.2; MgSO₄, 1.2; dextrose, 10.0; andNaHCO₃, 24.8. Tissue bath solutions were maintained at 37° C. andequilibrated with 95% O₂ and 5% CO₂. Tissues were placed under optimumresting force (4 g) and were allowed to equilibrate for approximately 1hour before exposure to the test compound. Isometric contractions wererecorded as changes in grams of force on a Beckman Dynograph withStatham UC-3 transducers.

Determination of Apparent Antagonist Dissociation Constant

Noncumulative contractile concentration-response curves for serotonin inthe fundus and cumulative concentration response curves in the jugularvein were obtained by a stepwise increase in concentration after washingout the preceding concentrations every 15-20 minutes. Each agonistconcentration remained in contact with the tissue for approximately 2minutes and maximum response to each compound concentration wasmeasured. ED₅₀ values were taken as the concentration of agonist thatproduced half-maximal contraction. After control responses wereobtained, tissues were incubated with an appropriate concentration ofbuffer or antagonist for 1 hour. Responses to serotonin were thenrepeated in the presence of an antagonist. Concentration responsesutilized only one agonist and one antagonist concentration per tissue.In general, successive agonist responses in the presence of buffertreatment were unaltered (average dose ratio was 1.28±0.21).

Apparent antagonist dissociation constants (K_(B)) were determined foreach concentration of antagonist according to the following equation:

    K.sub.B = B!/(dose ratio-1)

where B! is the concentration of the antagonist and dose ratio is theED₅₀ of the agonist in the presence of the antagonist divided by thecontrol ED₅₀. Generally, parallel shifts in the concentration-responsecurves occurred in the presence of antagonists. The results wereexpressed as the negative logarithm of the K_(B) (i.e., -log K_(B)).Calculations were completed using known methods. B. R. Zaborowsky, J.Pharmacol. Methods, 4:4165 (1980).

IP₃ Formation in 5-HT_(2B) Transformed Cells

Formation and Extraction of IP₃ : A600K-2-3-MTX cells, grown insuspension, were harvested by centrifugation at 200×g and wereresuspended in protein-free cell culture medium. After incubations ofthe cells (2.5-3×106 cells/tube in 125 μl) at 37° for 10 minutes, 125 μlof the compound of interest, diluted in protein-free medium, were added.All incubations were performed in triplicate. When antagonists were usedto inhibit the effect of 5-HT, the cells were incubated with theantagonists for 10 minutes at 37° before the addition of 5-HT. Afteraddition of agonist, the cell suspension was vortexed and incubated foran additional 10 seconds at 37° (the 10 seconds include the time forvortexing). Then 250 μl of ice-cold 10% perchloric acid were added toterminate the reaction. The tubes were incubated for 10 minutes on iceand then centrifuges at 1500×g for 10 minutes. After centrifugation, 400μl of the supernatant were sampled. The following IP₃ extractionprocedure was modified from published procedures (E. S. Sharps, et al, AHigh Performance Liquid Chromatographic Method To Measure ³² PIncorporation Into Phosphorylated Metabolites In Cultured Cells., Anal.Biochem. 124:421-424 (1982) and K. A. Wreggett, et al, A RapicSeparation Method For Inositol Phosphates And Their Isomers., Biochem.J., 245:655-660 (1987)). The 400 μl sample was added to a 1.5 mlmicrofuge tube containing 100 μl of 10 mM EDTA, pH 9.0. This wasfollowed by the addition of 500 μl of 1,1,2trichlorotrifluroethane/tri-n-octylamine (1:1, v/v). The tubes werevortexed vigorously for 5-7 minutes and then centrifuged at 1500×g for 2minutes to aid in separation of the three layers. From the top aqueouslayer 100 μl were sampled for the determination of IP₃ content by theassay described below.

IP₃ binding assay: Rat cerebellar membranes were used as the source forthe IP₃ -binding protein in a binding assay modified from publishedprocedures (P. F. Worley, et al, Characterization Of InositolTriphosphate Receptor binding in brain, J. Biol. Chem., 262:12132-12136(1987) and D. S. Bredt et al, A Simple, Sensitive, And SpecificRadioreceptor Assay For Inositol 1,4,5-Triphosphate In BiologicalTissues, Biochem. Biophys. Res. Commun., 159:976-982 (1989)). Membraneswere prepared by homogenizing rat cerebella in 30 volumes ofhomogenization buffer (1 mM EDTA and 1 mM 2-mercaptoethanol in 50 mMTris.HCl, pH 7.7), using a Tissumizer (Tekmar) at setting 65, for 15seconds. The homogenate was centrifuges at 39,800×g for 10 minutes at 4°. This procedure was repeated three more times, for a total of fourwashes. The final pellet was suspended in 30 volumes of IP₃ bindingbuffer (1 mM EDTA and 1 mM 2-mercaptoethanol in 64.3 mM Tris.HCl, pH9.0) and frozen at -70° until needed.

Binding buffer (350 μl, containing ³ H!IP₃ and 50 μl of binding proteinhomogenate were added to 100 μl of the extracted IP₃ samples or knownIP₃ standards that had been subjected to the extraction procedure asdescribed above. The final concentration of ³ H!IP₃ was 1 nM. The tubeswere incubated at 0° for 15 minutes and then filtered through WhatmanGF/B filters pre-wet with water and precooled with 2 ml of ice-cold IP₃wash buffer (1 mM EDTA in 50 mM Tris.HCl, pH 9.0)! by using a Brandelcell harvester. The filters were then rapidly washed two times with 1 mlof ice-cold IP₃ wash buffer. The amount of ³ H!IP₃ trapped on thefilters was determined by liquid scintillation counting. The amount ofIP₃ in the samples was determined by comparison with the standard curve.

When cells expressing the 5-HT_(2B) receptor were preincubated withmianserin, methysergide, rauwolscine, or 1-NP before the addition of5-HT, the 5-HT curves were shifted to the right and the E_(max) valueswere decreased, relative to 5-HT alone.

5-HT_(2A) and 5-HT_(2C) Receptor Binding Activity

The ability of a compound to bind to a 5-HT_(2A) or 5-HT_(2C) receptorwas measured using standard procedures such as that listed below.

Assay Procedure

Membrane preparation from transformed cell lines. Membranes wereprepared using AV12 cells (Syrian hamster fibroblast, ATCC no. CRL 9595)stably transformed with the human-5-HT_(2A), or 5-HT_(2C) receptor(Wainscott et al., Pharmacological Characteristics Of The Newly ClonedRat 5-Hydroxytryptamine_(2F) Receptor, Mol. Pharmacol., 48:419-426(1993)). Briefly, cells expressing the receptor of interest were grownin suspension and harvested by centrifugation. The cells wereresuspended in a minimal volume of a hypotonic buffer, 50 mM Tris-HCL,pH 7.4, and frozen at 70° C. until needed. On the day the assay, thesuspension was thawed and diluted to 35 ml/0.5×10² cells, original cellnumber, with 50 mM Tris-HCl, pH 7.4, and centrifuged at 39,800×g, 4° C.The resulting pellet was resuspended by vortexing and incubated at 37°C. for 10 min, then centrifuged at 39,800×g, 4° C. This pellet wasresuspended and centrifuged one more time. To achieve a homogenousmembrane suspension, the final pellet was resuspended using a Tissumizer(Tekmar, Cincinnati, Ohio) at setting 75 for 10 to 15 sec. in 67 mMTris-HCl, pH 7.4, for cells expressing the human or rat 5-HT_(2A)receptor or 67 mM Tris-HCl, pH 7.4, containing 13 mM MgCl₂ and 0.67 mMEDTA for cells expressing the human 5-HT_(2C) receptors.

5-HT_(2A),2C ¹²⁵ I!DOI binding studies: Human 5-HT_(2A) or 5-HT_(2C)binding studies were performed essentially as described for ³ H!5-HTbinding to the 5-HT_(2B) receptor with the following exceptions. Theassay buffer contained, in final concentration, 10 mM pargyline, 9.75 mMMgCl₂, 0.5 mM EDTA, 0.1% sodium ascorbate and 50 mM Tris-HCl, pH 7.4.Incubations were performed at 37° C. for 30 minutes with approximately40 and 30 mg of protein for the 5-HT_(2A) and 5-HT_(2C) receptors,respectively, then filtered through Whatman GF/C filters which had beenpresoaked in 0.5% (w/v) polyethylenimine and precooled with 4 ml ofice-cold wash buffer. The filters were then washed rapidly 4 times with1 ml of ice-cold wash buffer. The amount of ¹²⁵ I!DOI trapped on thefilters was determined using a gamma counter. Nonspecific binding wasdetermined with 10 mm mianserin for 5-HT_(2C) and 1 mM ketanserin for5-HT_(2A) receptors. The final concentration of ¹²⁵ I!DOI wasapproximately 0.07 to 0.15 mM for competition experiments.

Statistical analysis: Nonlinear regression analysis for the saturationand competition curves was performed as described previously (Wainscottet al., Pharmacological Characteristics Of The Newly Cloned Rat5-Hydroxytryptamine_(2F) Receptor, Mol. Pharmacol., 48:419-426 (1993)).One-way analysis of variance was performed on the pK₁ values (i.e., logK., molar) followed by the Tukay-Kramer Honestly Significant Differencetest (JMP; SAS Institute Inc., Cary, N.C.). IC₅₀ values from thecompetition curves were converted to K_(d) values using theCheng-Prusoff (1973) equation. For ¹²⁵ I!DOI-labeled receptors, theK_(d) of ¹²⁵ I!DOI for the 5-HT_(2A) or 5-HT_(2C) receptors wasdetermined using a rearrangement of the Cheng-Prusoff equation giving:K_(d) =IC 50- L!, where IC₅₀ is the concentration of unlabeled DOIcausing 50% inhibition of specific ¹²⁵ I!DOI binding and I!=the freeconcentration of ¹²⁵ I!DOI.

IP₃ Formation in 5-HT_(2A) and 5-HT_(2C) Transformed Cells

IP₃ formation assay in 5-HT_(2A) and 5-HT_(2C) transformed cells wasconducted in the same manner as IP₃ formation in 5-HT_(2B) transformedcells with the exception that human AHSlC-3S cells were used for5-HT_(2C) and human Hu2-3S cells were used for 5-HT_(2A).

The following Experiments are for testing the efficacy of the 5-HT₂antagonists for treating or ameliorating the symptoms of the common coldor allergic rhinitis.

Experiment #1

Wistar rats (250-350 g, Harlan Sprague Dawley, Indianapolis, Ind.) orHartley guinea pigs (250-350 g, Charles River Laboratories, Inc.,Wilmington, Mass.) are anesthetized with sodium pentobarbital (65 mg/kgor 45 mg/kg, respectively, intraperitoneally).

The femoral vein is exposed and a 50 mg/kg dose of Evans Blue, afluorescent dye, is injected intravenously (1 mL/kg). Approximately 2minutes later, a dose of mCPP, alpha-methyl serotonin, or saline is alsoinjected intravenously. The Evans Blue complexes with proteins in theblood and functions as a marker for protein extravasation. Exactly 15minutes post-injection of the agonist or saline, the animals are killedby exsanguination to rinse out the blood and dye left in the vessels.The posterior vena cava and the aorta of the animals are clamped and theright auricle is cut open to facilitate the process. The exsanguinationperfusion is performed by injection of saline (40 mL) into the leftventricle using an approximate infusion rate of 1 mL/sec.

The nasal membrane samples are removed from both sides, blotted dry on apaper towel, and weighed. The samples are then placed in individualtubes containing 3 ml of formamide and incubated at 37° C. for 18-24hrs. The formamide is separated from the tissue following the incubationand its optical density is read at 620 nm using a Beckmann Model OU-7spectrophotometer. The optical densities of three known concentrationsof Evans Blue dye are used to construct a standard curve from which theEvans Blue dye concentration of the unknown samples is determined. Allvalues are normalized by the weight of the original tissue sample.

Results

Intravenous injection of either mCPP or alpha-methyl serotonin producean increase in the amount of protein extravasation in the nasal tissueas indicated by the dye Evans Blue when compared to a saline control.

Experiment #2

Wistar rats (250-350 g, Harlan Sprague Dawley, Indianapolis, Ind.) orHartley guinea pigs (250-350 g, Charles River Laboratories, Inc.,Wilmington, Mass.) are anesthetized with sodium pentobarbital (65 mg/kgor 45 mg/kg, respectively, intraperitoneally).

The femoral vein is exposed and a 50 mg/kg dose of Evans Blue combinedwith a dose of either 6-methyl-1-(2-chloro-3,4-dimethoxyphenyl)-methyl!-1,2,3,4-tetrahydro-9H-pyrido3,4-b!indole hydrochloride, spiro-9,92-(3,4-dimethoxy)-1,2,3,4-tetrahydronaphthyl!-5-methyl-1,2,3,9-tetrahydro-8H-pyridoindole, 6-methyl-1-(2-bromo-3,4-dimethoxyphenyl)-methyl!-1,2,3,4-tetrahydro-9H-pyrido3,4-b!indole and 6-iodo-1-(3,4-dimethoxyphenyl)-methyl!-1,2,3,4-tetrahydro-9H-pyrido 3,4-b!indolehydrochloride is administered intravenously. Approximately 2 minuteslater, a dose of mCPP, alpha-methyl serotonin, or saline is alsoinjected intravenously. The Evans Blue complexes with proteins in theblood and functions as a marker for protein extravasation. Exactly 15minutes post-injection of the agonist or saline, the animals are killedby exsanguination to rinse out the blood and dye left in the vessels.The posterior vena cava and the aorta of the animals are clamped and theright auricle is cut open to facilitate the process. The exsanguinationperfusion is performed by injection of saline (40 mL) into the leftventricle using an approximate infusion rate of 1 mL/sec.

The nasal membrane samples are removed from both sides, blotted dry on apaper towel, and weighed. The samples are then placed in individualtubes containing 3 ml of formamide and incubated at 37° C. for 18-24hrs. The formamide is separated from the tissue following the incubationand its optical density is read at 620 nm using a Beckmann Model OU-7spectrophotometer. The optical densities of three known concentrationsof Evans Blue dye are used to construct a standard curve from which theEvans Blue dye concentration of the unknown samples is determined. Allvalues are normalized by the weight of the original tissue sample.

Results

Intravenous injection of either mCPP or alpha-methyl serotonin producean increase in the amount of protein extravasation in the nasal tissueas indicated by the dye Evans Blue when compared to a saline control.6-methyl-1-(2-chloro-3,4-dimethoxyphenyl)-methyl!-1,2,3,4-tetrahydro-9H-pyrido3,4-b!indole hydrochloride, spiro-9,92-(3,4-dimethoxy)-1,2,3,4-tetrahydronaphthyl!-5-methyl-1,2,3,9-tetrahydro-8H-pyridoindole, 6-methyl-1-(2-bromo-3,4-dimethoxyphenyl)-methyl!-1,2,3,4-tetrahydro-9H-pyrido3,4-b!indole and 6-iodo-1-(3,4-dimethoxyphenyl)-methyl!-1,2,3,4-tetrahydro-9H-pyrido 3,4-b!indolehydrochloride dose dependently inhibite the agonist-induced increases innasal tissue protein extravasation when administered 2 minutes prior toagonist challenge.

Formulations

While it is possible to administer a compound employed in the methods ofthis invention directly without any formulation, the compounds areusually administered in the form of pharmaceutical compositionscomprising a pharmaceutically acceptable excipient and at least oneactive ingredient (the compound of the present invention). Suchcompositions contain from about 0.1% by weight to about 90.0% by weightof the present compound. These compositions can be administered by avariety of routes including oral, rectal, transdermal, subcutaneous,intravenous, intramuscular, and intranasal. Many of the compoundsemployed in the methods of this invention are effective as bothinjectable and oral compositions. Such compositions are prepared in amanner well known in the pharmaceutical art and comprise at least oneactive compound. See, e.g., REMINGTON'S PHARMACEUTICAL SCIENCES, (16thed. 1980).

In making the compositions employed in the present invention the activeingredient is usually mixed with an excipient, diluted by an excipientor enclosed within such a carrier which can be in the form of a capsule,sachet, paper or other container. When the excipient serves as adiluent, it can be a solid, semi-solid, or liquid material, which actsas a vehicle, carrier or medium for the active ingredient. Thus, thecompositions can be in the form of tablets, pills, powders, lozenges,sachets, cachets, elixirs, suspensions, emulsions, solutions, syrups,aerosols (as a solid or in a liquid medium), ointments containing forexample up to 10% by weight of the active compound, soft and hardgelatin capsules, suppositories, sterile injectable solutions, andsterile packaged powders.

In preparing a formulation, it may be necessary to mill the activecompound to provide the appropriate particle size prior to combiningwith the other ingredients. If the active compound is substantiallyinsoluble, it ordinarily is milled to a particle size of less than 200mesh. If the active compound is substantially water soluble, theparticle size is normally adjusted by milling to provide a substantiallyuniform distribution in the formulation, e.g. about 40 mesh.

Some examples of suitable carriers, excipients and diluents includelactose, dextrose, sucrose, sorbitol, mannitol, starches, gum acacia,calcium phosphate, alginates, tragacanth, gelatin, calcium silicate,microcrystalline cellulose, polyvinylpyrrolidone, cellulose, tragacanth,gelatin, water, syrup, and methyl cellulose. The formulations canadditionally include: lubricating agents such as talc, magnesiumstearate, and mineral oil; wetting agents; emulsifying and suspendingagents; preserving agents such as methyl- and propylhydroxybenzoates;sweetening agents; and flavoring agents. The compositions of theinvention can be formulated so as to provide quick, sustained or delayedrelease of the active ingredient after administration to the patient byemploying procedures known in the art.

The compounds of this invention may be delivered transdermally usingknown transdermal delivery systems and excipients. Most preferably, acompound of this invention is admixed with permeation enhancersincluding, but not limited to, propylene glycol, polyethylene glycolmonolaurate, and azacycloalkan-2-ones, and incorporated into a patch orsimilar delivery system. Additional excipients including gelling agents,emulsifiers, and buffers may be added to the transdermal formulation asdesired.

For oral administration, a compound of this invention ideally can beadmixed with carriers and diluents and molded into tablets or enclosedin gelatin capsules.

The compositions are preferably formulated in a unit dosage form, eachdosage containing from about 0.05 to about 150 mg, more usually about1.0 to about 100 mg, of the active ingredient. The term "unit dosageform" refers to physically discrete units suitable as unitary dosagesfor human subjects and other mammals, each unit containing apredetermined quantity of active material calculated to produce thedesired therapeutic effect, in association with a suitablepharmaceutical excipient.

The active compounds are generally effective over a wide dosage range.For example, dosages per day normally fall within the range of about0.01 to about 30 mg/kg of body weight. In the treatment of adult humans,the range of about 0.1 to about 15 mg/kg/day, in single or divided dose,is especially preferred. However, it will be understood that the amountof the compound actually administered will be determined by a physician,in the light of the relevant circumstances, including the condition tobe treated, the chosen route of administration, the actual compound orcompounds administered, the age, weight, and response of the individualpatient, and the severity of the patient's symptoms, and therefore theabove dosage ranges are not intended to limit the scope of the inventionin any way. In some instances dosage levels below the lower limit of theaforesaid range may be more than adequate, while in other cases stilllarger doses may be employed without causing any harmful side effect,provided that such larger doses are first divided into several smallerdoses for administration throughout the day.

In order to more fully illustrate the operation of the presentinvention, the following formulation examples are provided. The examplesare illustrative only, and are not intended to limit the scope of theinvention. The formulations may employ as active ingredients (compounds)any of the compounds of the present invention.

Formulation Preparation 1

Hard gelatin capsules containing the following ingredients are prepared:

    ______________________________________                    Quantity    Ingredient      (mg/capsule)    ______________________________________    Active Ingredient(s)                    100.0    Starch          235.0    Magnesium stearate                    5.0    ______________________________________

The above ingredients are mixed and filled into hard gelatin capsules in340 mg quantities.

Formulation Preparation 2

A tablet formula is prepared using the ingredients below:

    ______________________________________                      Quantity    Ingredient        (mg/tablet)    ______________________________________    Active Ingredient(s)                      100.0    Cellulose, microcrystalline                      125.0    Colloidal silicon dioxide                      10.0    Stearic acid      5.0    ______________________________________

The components are blended and compressed to form tablets, each weighing240 mg.

Formulation Preparation 3

A dry powder inhaler formulation is prepared containing the followingcomponents:

    ______________________________________    Ingredient       Weight %    ______________________________________    Active Ingredient(s)                     5    Lactose          95    ______________________________________

The active mixture is mixed with the lactose and the mixture is added toa dry powder inhaling appliance.

Formulation Preparation 4

Tablets, each containing 30 mg of active ingredient, are prepared asfollows:

    ______________________________________                           Quantity    Ingredient             (mg/tablet)    ______________________________________    Active Ingredient(s)   30.0   mg    Starch                 45.0   mg    Microcrystalline cellulose                           35.0   mg    Polyvinylpyrrolidone   4.0    mg    (as 10% solution in water)    Sodium carboxymethyl starch                           4.5    mg    Magnesium stearate     0.5    mg    Talc                   1.0    mg    Total                  120    mg    ______________________________________

The active ingredient, starch and cellulose are passed through a No. 20mesh U.S. sieve and mixed thoroughly. The solution ofpolyvinylpyrrolidone is mixed with the resultant powders, which are thenpassed through a 16 mesh U.S. sieve. The granules so produced are driedat 50°-60° C. and passed through a 16 mesh U.S. sieve. The sodiumcarboxymethyl starch, magnesium stearate, and talc, previously passedthrough a No. 30 mesh U.S. sieve, are then added to the granules which,after mixing, are compressed on a tablet machine to yield tablets eachweighing 120 mg.

Formulation Preparation 5

Capsules, each containing 40 mg of medicament are made as follows:

    ______________________________________                          Quantity    Ingredient            (mg/capsule)    ______________________________________    Active Ingredient(s)  40.0   mg    Starch                109.0  mg    Magnesium stearate    1.0    mg    Total                 150.0  mg    ______________________________________

The active ingredient, cellulose, starch, and magnesium stearate areblended, passed through a No. 20 mesh U.S. sieve, and filled into hardgelatin capsules in 150 mg quantities.

Formulation Preparation 6

Suppositories, each containing 25 mg of active ingredient are made asfollows:

    ______________________________________    Ingredient              Amount    ______________________________________    Active Ingredient(s)    25     mg    Saturated fatty acid glycerides to                            2,000  mg    ______________________________________

The active ingredient(s) is passed through a No. 60 mesh U.S. sieve andsuspended in the saturated fatty acid glycerides previously melted usingthe minimum heat necessary. The mixture is then poured into asuppository mold of nominal 2.0 g capacity and allowed to cool.

Formulation Preparation 7

Suspensions, each containing 50 mg of medicament per 5.0 ml dose aremade as follows:

    ______________________________________    Ingredient               Amount    ______________________________________    Active Ingredient(s)     50.0   mg    Xanthan gum              4.0    mg    Sodium carboxymethyl cellulose (11%)                             50.0   mg    Microcrystalline cellulose (89%)    Sucrose                  1.75   g    Sodium benzoate          10.0   mg    Flavor and Color         q.     v.    Purified water to        5.0    ml    ______________________________________

The medicament, sucrose and xanthan gum are blended, passed through aNo. 10 mesh U.S. sieve, and then mixed with a previously made solutionof the microcrystalline cellulose and sodium carboxymethyl cellulose inwater. The sodium benzoate, flavor, and color are diluted with some ofthe water and added with stirring. Sufficient water is then added toproduce the required volume.

Formulation Preparation 8

Capsules, each containing 15 mg of medicament, are made as follows:

    ______________________________________                          Quantity    Ingredient            (mg/capsule)    ______________________________________    Active Ingredient(s)  15.0   mg    Starch                407.0  mg    Magnesium stearate    3.0    mg    Total                 425.0  mg    ______________________________________

The active ingredient(s), cellulose, starch, and magnesium stearate areblended, passed through a No. 20 mesh U.S. sieve, and filled into hardgelatin capsules in 425 mg quantities.

Formulation Preparation 9

An intravenous formulation may be prepared as follows:

    ______________________________________    Ingredient            Quantity    ______________________________________    Active Ingredient(s)  250.0  mg    Isotonic saline       1000   ml    ______________________________________

Formulation Preparation 10

A topical formulation may be prepared as follows:

    ______________________________________    Ingredient            Quantity    ______________________________________    Active Ingredient(s)  1-10    g    Emulsifying Wax       30      g    Liquid Paraffin       20      g    White Soft Paraffin   to 100  g    ______________________________________

The white soft paraffin is heated until molten. The liquid paraffin andemulsifying wax are incorporated and stirred until dissolved. The activeingredient is added and stirring is continued until dispersed. Themixture is then cooled until solid.

Formulation Preparation 11

Sublingual or buccal tablets, each containing 10 mg of activeingredient, may be prepared as follows:

    ______________________________________                           Quantity    Ingredient             Per Tablet    ______________________________________    Active Ingredient(s)   10.0   mg    Glycerol               210.5  mg    Water                  143.0  mg    Sodium Citrate         4.5    mg    Polyvinyl Alcohol      26.5   mg    Polyvinylpyrrolidone   15.5   mg    Total                  410.0  mg    ______________________________________

The glycerol, water, sodium citrate, polyvinyl alcohol, andpolyvinylpyrrolidone are admixed together by continuous stirring andmaintaining the temperature at about 90° C. When the polymers have goneinto solution, the solution is cooled to about 50°-55° C. and themedicament is slowly admixed. The homogenous mixture is poured intoforms made of an inert material to produce a drug-containing diffusionmatrix having a thickness of about 2-4 mm. This diffusion matrix is thencut to form individual tablets having the appropriate size.

Another preferred formulation employed in the methods of the presentinvention employs transdermal delivery devices ("patches"). Suchtransdermal patches may be used to provide continuous or discontinuousinfusion of the compounds of the present invention in controlledamounts. The construction and use of transdermal patches for thedelivery of pharmaceutical agents is well known in the art. See, e.g.,U.S. Pat. No. 5,023,252, issued Jun. 11, 1991, herein incorporated byreference. Such patches may be constructed for continuous, pulsatile, oron demand delivery of pharmaceutical agents.

Indirect techniques, which are generally preferred, usually involveformulating the compositions to provide for drug latentiation by theconversion of hydrophilic drugs into lipid-soluble drugs or prodrugs.Latentiation is generally achieved through blocking of the hydroxy,carbonyl, sulfate, and primary amine groups present on the drug torender the drug more lipid soluble and amenable to transportation acrossthe blood-brain barrier. Alternatively, the delivery of hydrophilicdrugs may be enhanced by intra-arterial infusion of hypertonic solutionswhich can transiently open the blood-brain barrier.

The type of formulation employed for the administration of the compoundsemployed in the methods of the present invention may be dictated by theparticular compounds employed, the type of pharmacokinetic profiledesired from the route of administration and the compound(s), and thestate of the patient.

What is claimed is:
 1. A method for treating or ameliorating thesymptoms of the common cold or allergic rhinitis in a mammal whichcomprises administering to the mammal an effective amount of a compoundor composition having activity as a 5-HT_(2B) receptor antagonist.
 2. Amethod for treating or ameliorating the symptoms of the common cold orallergic rhinitis in a mammal which comprises administering to themammal an effective amount of a compound of the formula: ##STR20##wherein R¹ and R³, independently, are hydrogen or C₁ -C₃ alkyl;n and n',independently, are 1, 2, or 3; R⁶ is hydrogen, C₁ -C₆ alkyl, C₂ -C₆alkenyl, halo, halo(C₁ -C₆)alkyl, halo(C₂ -C₆)alkenyl, COR⁵, C₁ -C₁₀alkanoyl, CO₂ R⁵, (C₁ -C₆ alkyl)_(m) amino, NO₂, --SR⁵, or OR⁵ ; R⁷ andR⁸, independently, are an R⁶ group or C₇ -C₁₆ arylalkyl; m is 1 or 2; R⁵is hydrogen or C₁ -C₄ alkyl; R⁵ is C₁ -C₄ alkyl; and --is optionally abond; or a pharmaceutically acceptable salt or solvate thereof.
 3. Amethod for treating or ameliorating the symptoms of the common cold orallergic rhinitis in a mammal which comprises administering to themammal an effective amount of a compound of the formula: ##STR21##wherein Q is hydrogen or (CHR₂)R₄ ;R₁, R₂ and R₃, independently, arehydrogen or C₁ -C₃ alkyl; R₄ is C₅ -C₈ cycloalkyl, substituted C₅ -C₈cycloalkyl, C₅ -C₈ cycloalkenyl, substituted C₅ -C₈ cycloalkenyl, or abicyclic or substituted bicyclic group; A is ##STR22## wherein R₆ andR₇, independently, are hydrogen, C₁ -C₆ alkyl, C₂ -C₆ alkenyl, halo,halo(C₁ -C₆)alkyl, halo(C₂ -C₆)alkenyl, COR₅, C₁ -C₁₀ alkanoyl, CO₂ R₅,(C₁ -C₆ alkyl)_(m) amino, NO₂, --SR₅, or OR₅ ; or R₆ and R₇ togetherwith the carbon atoms of group A form a 5-to 8-member carbon ring; m is1 or 2; R₅ is hydrogen or C₁ -C₄ alkyl; R₅, is C₁ -C₄ alkyl; and R₈ isan R₆ group, C₃ -C₈ cycloalkyl, substituted C₃ -C₈ cycloalkyl, C₃ -C₈cycloalkyl-(C₁ -C₃)alkyl, C₅ -C₈ cycloalkenyl, substituted C₅ -C₈cycloalkenyl, C₅ -C₈ cycloalkenyl-(C₁ -C₃)alkyl, or C₇ -C₁₆ arylalkyl;ora pharmaceutically acceptable salt or solvate thereof.
 4. The method ofclaim 3 wherein R₁ and R₃ are hydrogen, ##STR23## A is and Q iscyclohexylmethyl.
 5. A method for treating or ameliorating the symptomsof the common cold or allergic rhinitis in a mammal which comprisesadministering to the mammal an effective amount of a compound of theformula: ##STR24## wherein x is CO, CS or CH₂ ;R is R¹, YR¹, NR² R³ orOC(O)R¹ ; R¹ is hydrogen, C₁ -C₂₄ alkyl, C₂ -C₂₄ alkenyl, C₃ -C₈cycloalkyl, C₃ -C₈ cycloalkenyl or C₄ -C₃₂ cycloalk(en)ylalk(en)yl,optionally substituted with one or two hydroxy groups, or phenyloptionally substituted with one or more halogen, trifluoromethyl, C₁ -C₄alkyl, C₁ -C₄ alkoxy, C₁ -C₄ alkylthio, acyloxy, or cyano substituents;Y is O or S; R² and R³ independently are an R¹ group, or R² and R₃combined together form a four to eight member heterocyclic ringcontaining from one to three nitrogen atoms and from zero to threeoxygen or sulfur atoms;provided that, 1) R can only be OC(O)R¹ when X isCH₂, and 2) R can only be R¹ when X is CO or CS; or a pharmaceuticallyacceptable salt thereof.
 6. A method for treating or ameliorating thesymptoms of the common cold or allergic rhinitis in a mammal whichcomprises administering to the mammal an effective amount of a compoundof the formula: ##STR25##
 7. A method for treating or ameliorating thesymptoms of the common cold or allergic rhinitis in a mammal whichcomprises administering to the mammal an effective amount of a compoundof the formula: ##STR26## wherein Ar is phenyl; phenyl having at leastone halogen, lower alkyl, lower alkoxy, hydroxy, trifluoromethyl, orcyano substituent; or a heteroaromatic group selected from 2-thienyl,3-thienyl, 2-furanyl, 3-furanyl, 2-oxazolyl, 2-imidazolyl, 2-pyridyl,3-pyridyl, and 4-pyridyl;the dotted line is an optional double bond; Xand X¹ independently are hydrogen, halogen, lower alkyl, lower alkoxy,hydroxy, lower alkylthio, lower alkylsulfonyl, lower alkylamino, lowerdialkylamino, cyano, trifluoromethyl, or trifluoromethylthio; or X andX¹ taken together form a 5 to 7 membered carbocyclic ring; R¹ ishydrogen, lower alkyl or alkyl substituted with one or two hydroxygroups; R is a substituent having the formula: ##STR27## wherein n is aninteger from 2-6 inclusive; W is oxygen or sulfur; V¹ is OR⁴, SR⁵, CHR⁶R⁷, or NR⁸ R₉ ; and R³ to R⁹ independently are hydrogen, lower alkyl,lower alkenyl, cycloalkyl, or lower alkyl or lower alkenyl substitutedwith one or two hydroxyl groups;provided that: when X is hydrogen orfluoro, R¹ cannot be hydrogen; or a pharmaceutically acceptable acidaddition salt thereof.
 8. A method for treating or ameliorating thesymptoms of the common cold or allergic rhinitis in a mammal whichcomprises administering to the mammal an effective amount of a compoundof the formula: ##STR28## wherein R₁, R₂ and R₃ independently arehydrogen or C₁ -C₆ alkyl;X is hydrogen or halogen; Z is a carbonyl ormethylene group; and C₉ ------C₁₀ represents a single or a doublebond;or a racemate or acid additional salt thereof.